A Conversation on Aging Theories

A month or so ago, I posted some of Robert Bradbury's thoughts on a grand unified theory of aging. A few days ago, Joao Magalhaes - a microbiologist and aging researcher - offered some comments:

My response time is a bit slower than usual but I thought I have a go at this nonetheless.
At 18:49 03-04-2004, you wrote:

First, lets assume the Free Radical theory of Aging which involves various aspects of Mitochondrial damage and aging are correct. [This explains why caloric restriction works.]

### Actually, I have a paragraph in a manuscript I'm working on that may interest you:

"Given the large number of age-related changes, it is crucial to evaluate the predictions of each theory and follow a system-level approach of which genetic manipulations in animal models are a crucial tool. The mechanisms of CR are a perfect example for the open-minded scepticism that should be implemented in ageing research. As mentioned earlier, both the GH/IGF-1 axis and ROS production appear to change in CR. Hypothesis aiming to explain CR based on either mechanism are theoretically sound. Yet while a clear cause-effect phenotype is witnessed when manipulating the components of the GH/IGF-1 axis, such is not the case for ROS. Therefore, the GH/IGF-1 axis, not oxidative damage, is the main candidate for explaining CR."

By manipulating the components I mean genetic manipulations in animal models, such as mice. If you genetically manipulate a number of components of the GH/IGF-1 axis you get a phenotype very similar to CR. If you manipulate ROS production or damage, you don't. So a clear cause-effect phenotype is witnessed suggesting the GH/IGF-1 axis is, at least partly, the mechanism involved in CR. No direct evidence exists that ROS are a cause of CR.

Second, lets assume you can't do too much about them because radicals and/or other pro-oxidants (e.g. nitric oxide) are being used as signal molecules (this may be somewhat controversial).

### I agree with the view that ROS are signalling molecules and there is good evidence supporting this view and supporting the idea that ROS are important in development.

Third, lets assume that the free radicals lead to DNA mutations (which is one way cancer develops) or worse leads to DNA double strand breaks. (Radiation and perhaps toxic substances in food or the environment might contribute to this as well).

### There isn't any direct evidence that free radical damage to the DNA causes ageing in mammals. Again, a paragraph from the same manuscript:

"One possibility is that ROS damage DNA and some evidence exists showing an increase in oxidative damage to DNA with age (Hamilton et al., 2001). Yet overexpression of catalase in the nucleus did not prevent oxidative damage to DNA (Schriner et al., 2000) and knocking out the gene responsible for 8-oxo-dGTPase failed to accelerate ageing (Tsuzuki et al., 2001). These results hint that the free radical and the DNA damage theories of ageing are not complementary."

BTW, 8-oxo-dGTPase repairs 8-oxo-7,8-dihydroguanine, an abundant and mutagenic form of oxidative DNA damage.

What I'm saying is that although oxidative damage to the DNA accumulates with age in some tissues, it is not proven this is a cause rather an effect of ageing. And the cited experiments suggest this is an effect of ageing.

Refs:

Hamilton, M.L., Van Remmen, H., Drake, J.A., Yang, H., Guo, Z.M., Kewitt, K., Walter, C.A., Richardson, A., 2001. Does oxidative damage to DNA increase with age? Proc Natl Acad Sci U S A 98, 10469-10474.

Schriner, S.E., Ogburn, C.E., Smith, A.C., Newcomb, T.G., Ladiges, W.C., Dolle, M.E., Vijg, J., Fukuchi, K., Martin, G.M., 2000. Levels of dna damage are unaltered in mice overexpressing human catalase in nuclei. Free Radic Biol Med 29, 664-673.

Tsuzuki, T., Egashira, A., Igarashi, H., Iwakuma, T., Nakatsuru, Y., Tominaga, Y., Kawate, H., Nakao, K., Nakamura, K., Ide, F., Kura, S., Nakabeppu, Y., Katsuki, M., Ishikawa, T., Sekiguchi, M., 2001. Spontaneous tumorigenesis in mice defective in the MTH1 gene encoding 8-oxo-dGTPase. Proc Natl Acad Sci U S A 98, 11456-11461.

As for the rest of your theory, Robert, I agree with the view that changes in DNA over time play a role in ageing. This could even be the case in stem cell stocks, as suggested by Van Zant's work. Yet I have some reservations on the idea that only one type of DNA damage causes ageing. At a first glance, it just doesn't seem right. Like I wrote previously on the mechanisms of CR, your idea is theoretically sound but there is no direct evidence in support of it. Can you think of one or two experiments that can prove or disprove your theory?

Robert Bradbury replied with:

What I'm saying is that although oxidative damage to the DNA accumulates with age in some tissues, it is not proven this is a cause rather an effect of ageing. And the cited experiments suggest this is an effect of ageing.

Well (Van Remmen, 2003 [1]) is interesting in that free radicals seem to be contributing to cancer but not to ageing. Though the standard caveat should be added that this is research in mice and not in humans.

In this case the entire somatic mutation approach may need to be modified such that it is only involved in cancer -- which is the cause of death for ~30% of people.

As for the rest of your theory, Robert, I agree with the view that changes in DNA over time play a role in ageing. This could even be the case in stem cell stocks, as suggested by Van Zant's work. Yet I have some reservations on the idea that only one type of DNA damage causes ageing. At a first glance, it just doesn't seem right. Like I wrote previously on the mechanisms of CR, your idea is theoretically sound but there is no direct evidence in support of it. Can you think of one or two experiments that can prove or disprove your theory?

As Aubrey pointed out to me the theory as described ignores possible epigenetic changes. Which in light of the difference between the progeria gene and the W.S. gene functions may be of significant concern. [See [2] for example when causes/symptoms of the two may be difficult to separate.]

I'm perfectly willing to accept that stem cell depletion may play an important role in aging -- that would sound like one needs an experiment to measure apoptosis (human cells are very intolerant of double strand breaks and might undergo apoptosis at a higher rate than mice) and replacement by stem cells with age. The recent report by (Li, 2004 [3]) suggests that you cannot separate Werner's Syndrome type aging from a direct involvement with double strand breaks (which is what PARP is involved in dealing with). It sounds like we need some kind of complex pulse-chase experiment with long-lived radioisotopes to measure cell death and stem cell replacement rates.

Now, with regard to the double strand break problem itself -- the problem in my mind is *where* do the breaks occur? I think you can get at this problem with high amplification PCR. If you assume the breaks are taking place at random places in the genome (but perhaps with some bias with respect to junk DNA, regulatory DNA and expressed DNA) then if you have enough cells and simply amplify various regions of the genome you should be able to gather statistical evidence with respect to where breaks are taking place and how they may impact gene expression. This may feedback into the actual structure of DNA in the nucleus and the whole LAMIN/progeria/epigenetic story.

If the DSB somatic mutation theory is correct you should be able to pull out the locations where breaks (and microdeletions) have taken place by differences in the length of the PCR amplification products. Vijg's (and others) work suggests that the deletions are taking place but to the best of my knowledge nobody has tried to do a statistical analysis of where they occur directly.

Finally, one needs mutant mice knockouts in at least the W.S. gene as well as the Artemis gene (perhaps heterozygous) to see if that impacts the frequency of microdeletions significantly.

1. Van Remmen H, Ikeno Y, Hamilton M, Pahlavani M, Wolf N, Thorpe SR, Alderson NL, Baynes JW, Epstein CJ, Huang TT, Nelson J, Strong R, Richardson A.Physiol Genomics. 2003 Dec 16;16(1):29-37. Life-long reduction in MnSOD activity results in increased DNA damage and higher incidence of cancer but does not accelerate aging. mutations in atypical Werner's syndrome.

2. Chen L, Lee L, Kudlow BA, Dos Santos HG, Sletvold O, Shafeghati Y, Botha EG, Garg A, Hanson NB, Martin GM, Mian IS, Kennedy BK, Oshima J. LMNA mutations in atypical Werner's syndrome. Lancet. 2003 Aug 9;362(9382):440-5

3. Li B, Navarro S, Kasahara N, Comai L. J Biol Chem. 2004 Apr 2;279(14):13659-67. Epub 2004 Jan 20. Identification and biochemical characterization of a Werner's syndrome protein complex with Ku70/80 and poly(ADP-ribose) polymerase-1.

Finally, Joao commented:

As with all theories of ageing, my major problem is how to prove that the mechanism you propose is a cause rather than an effect of ageing. For instance, you place a lot of weight on double strand breaks (DSB) as a causal factor in ageing. My advice would be for you to make a list of proteins involved in DSB repair and the phenotype witnessed in mice or humans when these proteins are mutated. At the IABG10 meeting in Cambridge last September, Vijg gave a talk along these lines in considering DSB repair mechanisms as critical in ageing but I was left mostly unconvinced. He did not show a clear trend in mutations affecting DSB influencing ageing. For instance, don't p53 mutations affect DSB? What about Pms2? Both these mutations decrease the lifespan of mammals but don't appear to change ageing. Nibrin also affects DSB in humans and yet mutations in NSB1 do not appear to accelerate ageing--at least not to me, though Vijg has argued it does.

The manuscript I'm working on deals exactly with the issue of segregating cause and effect of ageing. Animal models, such as mice, and human progeroid syndromes offer the best evidence of causal mechanisms in ageing. My opinion is that these are the best set of observations we have for understanding ageing and by fitting these observations together we may be able to develop an holistic view of the mechanisms of ageing. I'll warn you when I have a final version of the manuscript but it looks good.

Interesting stuff, isn't it? If you were subscribed to the Extropy-Chat mailing list, you'd see this sort of science discussed fairly often.

Comments

Keep one thing in mind when reading discussions of factors that cause aging: Much of what we need to develop as capabilities to repair and rejuvenated aged bodies are independent of of which mechanisms end up being most important in causing aging much.

Think about this intuitively. If your car's alternator breaks you need a new one. It doesn't matter why it breaks. We can fix the car because we can replace the alternator. The same is true of car parts in general. One breaks and we fix or replace it.

Another important point to note about repair and rejuvenations therapies is that if we can develop and apply therapies that repair or replace cells or organs we will be able to learn how important various causes of aging are just by seeing how much various repair techniques help. The point here is that the development of repair therapies will provide both useful therapies and tools to use to understand the causes of aging.

We can take an engineering approach to aging and start developing and trying out rejuvenation therapies on animal models as a way to both test therapies and to better understand the science of aging. The engineering approach does not have to wait on advances in the science of aging mechanisms. We can do much of the bioengineering first without waiting for the science to explain why some approaches to treatment work better than other approaches.

Posted by: Randall Parker at April 20th, 2004 2:39 PM

Aubrey de Grey added:

> Well (Van Remmen, 2003 [1]) is interesting in that free radicals seem
> to be contributing to cancer but not to ageing.

Nearly but not quite -- what it shows is that free radicals accessible to antioxidant enzymes do not contribute to aging. (and, as you say, this is only in lab mice, not humans and not even wild mice.) This is important because there is a distinct possibility that superoxide does most of its harm before it reaches a location where SOD can get at it. I suggested in 2000 (JAAM 3:25) that the action might be occurring in the intermembrane space, where superoxide might be protonated rather more often than elsewhere (more acidic because of proton-pumping by oxidative phosphorylation). The protonated form of superoxide is a really nasty species (see my subsequent paper DNA Cell Biol 21:251), in contrast to the tameness of superoxide itself. There are actually a few weakneses in my proposal, but they were neatly eliminated by an idea proposed by Florian Muller a few months later (J Am Aging Assoc 23:227): he looked hard at the structure of Complex III and realised that protonated superoxide could be formed within the actual membrane itself (in the lipid phase), and zap a fatty acid or protein at once. Coincidentally Muller is now a coworker of Van Remmen in San Antonio.

Posted by: Reason at April 22nd, 2004 10:32 PM

First, I believe the cause of aging will also ultimately be the body's repair.

The posted statements and observations clearly point to lack of hormonal communication as the deity of aging (i.e. its cause). However, a much more complex, but clear, picture is drawn out for us by Dr. Barry Sears in the Anti-Aging Zone pointing to multiple pillars of aging all of which explains why calorie restriction works at each levels.

Posted by: Scott Fabyanic at June 6th, 2004 3:56 PM

Mitochondria damage is not the cause of aging it is one of the many consequences. It is clear that calorie restriciotn works on all levels in cluding reducing that damage. Most importantly it reduces silent inflammation (excess amounts of Arachadonic Acid) which is the underlying CAUSE of all chronic disease and the aging process itself. Oxidation and Glycation take distance "thirds". Therefore, the reduction of excess' silent inflammation becomes the Holy Grail of Anti-Aging not to mention the vast reduction of all Chronic Disease. This is best accomplished by Super, High Dose, High-Grade Fish Oils (EPA/DHA?Sesamol/GLA) and a moderate balanced restricted diet (40/30/30).

Posted by: Scott Fabyanic at July 8th, 2007 11:36 PM

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